[Metabolomic interference induced by short-chain chlorinated paraffins in human normal hepatic cells].

Short-chain chlorinated paraffins (SCCPs) are an emerging class of persistent organic pollutants (POPs) that are widely detected in environmental matrices and human samples. Because of their environmental persistence, long-range transport potential, bioaccumulation potential, and biotoxicity, SCCPs pose a significant threat to human health. In this study, metabolomics technology was applied to reveal the metabolomic interference in human normal hepatic (L02) cells after exposure to low (1 µg/L), moderate (10 µg/L), and high (100 µg/L) doses of SCCPs. Principal component analysis (PCA) and metabolic effect level index (MELI) values showed that all three SCCP doses caused notable metabolic perturbations in L02 cells. A total of 72 metabolites that were annotated by MS/MS and matched with the experimental spectra in the Human Metabolome Database (HMDB) or validated by commercially available standards were selected as differential metabolites (DMs) across all groups. The low-dose exposure group shared 33 and 36 DMs with the moderate- and high-dose exposure groups, respectively. The moderate-dose exposure group shared 46 DMs with the high-dose exposure group. In addition, 33 DMs were shared among the three exposure groups. Among the 72 DMs, 9, 9, and 45 metabolites participated in the amino acid, nucleotide, and lipid metabolism pathways, respectively. The results of pathway enrichment analysis showed that the most relevant metabolic pathways affected by SCCPs were the lipid metabolism, fatty acid ß-oxidation, and nucleotide metabolism pathways, and that compared with low-dose exposure, moderate- and high-dose SCCP exposures caused more notable perturbations of these metabolic pathways in L02 cells. Exposure to SCCPs perturbed glycerophospholipid and sphingolipid metabolism. Significant alterations in the levels of phosphatidylcholines, phosphatidylethanolamines, and sphingomyelins indicated SCCP-induced biomembrane damage. SCCPs inhibited fatty acid ß-oxidation by decreasing the levels of short- and medium-chain acylcarnitines in L02 cells, indicating that the energy supplied by fatty acid oxidation was reduced in these cells. Furthermore, compared with low- and moderate-dose SCCPs, high-dose SCCPs produced a significantly stronger inhibition of fatty acid ß-oxidation. In addition, SCCPs perturbed nucleotide metabolism. The higher hypoxanthine levels observed in L02 cells after SCCP exposures indicate that SCCPs may induce several adverse effects, including hypoxia, reactive oxygen species production, and mutagenesis in L02 cells.

Effects of Saccharomyces cerevisiae hydrolysate on growth performance, immunity function, and intestinal health in broilers.

The current study was performed to explore the effects of dietary supplementation of Saccharomyces cerevisiae hydrolysate (SCH) on growth performance, immune function, and intestinal health in broiler chicken. A total of 300 Ross 308 male broilers (1-day-old) were randomly assigned to 2 dietary treatments including a basal diet (control group), and a basal diet supplemented with SCH feed additive (500 mg/kg in starter and grower phase, and 250 mg/kg in finisher phase). Each treatment had 6 replicates with 25 birds each. The results showed that the addition of SCH promoted growth during d 15 to 28 (P < 0.05). Although the addition of SCH had no significant effect on the intestinal relative indexes, it significantly increased the jejunum villus height (VH) and the ratio of villus height to crypt depth (VCR) of jejunum, and decreased the crypt depth (CD) of ileum (P < 0.05). Furthermore, SCH addition significantly downregulated the mRNA expression of immunomodulatory genes (TNF-α, IL-1ß, and IL-6), and upregulated the tight junction genes (ZO-1 and Claudin-1) (P < 0.05). High throughput sequencing analysis of bacterial 16S rRNA revealed that dietary SCH supplementation altered cecum microbiota. Alpha diversity analysis showed that a higher bacterial richness in cecum of broilers fed with SCH. The composition of cecum microbiota regulated by SCH addition was characterized by an increased abundance of Firmicutes and a reduced abundance of Bacteroidetes. At the genus level, dietary SCH resulted in a decrease of Bacteroides and an increase of short-chain fatty acids (SCFA) -producing bacteria including Lactobacillus and Faecalibacterium. Taken together, dietary SCH supplementation can stimulate the growth of broilers by regulating the intestinal immunity and barrier function, and improving the intestinal morphology, which may be related to the enhancement of bacterial diversity and the changes of intestinal microbial composition.

Effects of Dietary Selenium Sources on Physiological Status of Laying Hens and Production of Selenium-Enriched Eggs.

Developing new sources of organic selenium (Se) has potential benefits for animal production and human nutrition via animal-based foods enriched with Se. The objective of this study was to evaluate the effects of Se-enriched insect protein (SEIP) in comparison with other sources, such as sodium selenite (SS) and selenium-enriched yeast (SEY), on performance, egg quality, selenium concentration in eggs, serum biochemical indices, immune capacity, and intestinal morphology of laying hens. Four hundred and fifty 24-week-old Hy-Line Brown laying hens with 94.0 ± 1.5% laying rate were randomly allocated to five groups with six replicates of 15 hens each. The control diet was prepared without adding exogenous selenium (calculated basal Se content of 0.08 mg/kg). The normal group was fed basal diets supplemented with 0.3 mg/kg of Se provided by sodium selenite. Three treatment groups (SS, SEY, and SEIP, respectively) were fed basal diets supplemented with 2 mg/kg of Se provided by sodium selenite, Se-enriched yeast, and SEIP, respectively. The feeding trial lasted for 12 weeks. Results revealed that dietary supplementation of 2 mg/kg of Se increased egg weight, decreased feed conversion ratio, and enhanced the antioxidant capacity of eggs in laying hens relative to the control group, whereas no significant differences were observed among SS, SEY, and SEIP treatment groups for the same. The organic source of Se provided by SEY or SEIP showed higher bio efficiency, as indicated by higher selenium content in eggs of SEY and SEIP compared with SS, although higher content was observed in SEY compared with SEIP. Also, the organic Se source significantly improved antioxidant capacity and immune functions of laying hens than the inorganic Se source. Diets supplemented with SEIP and SS significantly improved jejunal morphology of the laying hens compared with SEY, whereas SEIP was more effective than SEY to improve the oviduct health of laying hens. The results of this work evidently points the additive effect and nontoxicity of SEIP. Thus, SEIP could be used as another organic source of Se in the diet of laying hens and production of selenium-enriched eggs for humans.

Supplemental L-Arginine Improves the Embryonic Intestine Development and Microbial Succession in a Chick Embryo Model.

Early colonization of intestinal microbiota plays an important role in intestinal development. However, the microbial succession at an embryonic stage and its assembly patterns induced by prenatal nutrition are unknown. In the present study, we used a chick embryo model to investigate the effects of in ovo feeding (IOF) of L-arginine (Arg) on the intestinal development and microbial succession of embryos. A total of 216 fertile eggs were randomly distributed into 2 groups including the non-injected control group and IOF of Arg group with 7 mg/egg. The results showed that IOF Arg increased the intestinal index, absolute weight of jejunum, and improved jejunal morphology in terms of villus width and surface area (p < 0.05). The relative mRNA expressions of mTOR and 4E-BP1 were up-regulated and accompanied by higher contents of Mucin-2 in the Arg group (p < 0.05). There was a significant elevation in contents of serum glucose and high-density lipoprotein cholesterol, whereas there was a decreased low-density lipoprotein cholesterol in the Arg group (p < 0.05). Additionally, Proteobacteria and Firmicutes were major intestinal bacteria species at the embryonic stage. However, Arg supplementation targeted to shape assembly patterns of microbial succession and then changed microbial composition (p = 0.05). Meanwhile, several short-chain fatty acids (SCFAs)-producing bacteria, such as Roseburia, Blautia, and Ruminococcus were identified as biomarkers in the Arg group (LDA > 3, p < 0.05). Accordingly, significant elevated concentrations of SCFAs, including lactic acid and formic acid, were observed in the Arg group (p < 0.05), accompanied by the higher concentration of butyric acid (0.05 < p < 0.10). In conclusion, prenatal Arg supplementation improved embryonic intestine development by regulating glucose and lipid homeostasis to supply more energy for chick embryos. The possible mechanism could be the roles of Arg in shaping the microbial assembly pattern and succession of the embryonic intestine, particularly the enrichment of potential probiotics. These findings may contribute to exploring nutritional strategies to establish health-promoting microbiota by manipulating prenatal host-microbe interactions for the healthy development of neonates.

Effect of dietary serine supplementation on performance, egg quality, serum indices, and ileal mucosal immunity in laying hens fed a low crude protein diet.

This study was designed to evaluate the effect of dietary Ser on performance, egg quality, serum indices, and ileal mucosal immunity in laying hens fed low crude protein (LCP), essential amino acids (EAA) balanced diets. A total of 480 Hy-Line Brown layers at 24 wk of age were randomly assigned into 5 dietary treatments with 8 replicates of 12 birds each. Treatments included a control diet (16.49% CP), and 4 LCP, EAA balanced diets (14.05% CP) supplemented with 0, 0.114%, 0.306%, 0.498% L-Ser, respectively. Dietary Ser supplementation linearly increased hen-day egg production (HDEP; P < 0.05) and decreased feed-to-egg ratio (P < 0.05) among LCP groups from wk 6 to 10, and the optimal HDEP of layers occurred at Ser level of 0.498%. At the end of wk 10, birds in the control had higher albumen height and thick white proportion than those fed the LCP diet without Ser addition (P < 0.05), and presented a lower yolk color score than all LCP groups (P < 0.05). Among LCP groups, serum total protein and globulin contents were significantly increased by dietary Ser addition at the levels of 0.306% and 0.498% (P < 0.05), and had a linear response to the supplemental Ser levels (P < 0.05). Furthermore, dietary 0.498% Ser supplementation significantly increased serum immunoglobulin G and immunoglobulin M contents (P < 0.05) and up-regulated the expression of mucin 2, secretory immunoglobulin A, and relevant glycosyltransferases of O-glycosylation in ileal mucosa (P < 0.05). The increased expression of proinflammatory cytokines IFN-γ and IL-1ß induced by LCP diets (P < 0.05) was reversed following 0.498% Ser addition (P < 0.05). Collectively, dietary CP reduction by 2.44% could maintain the productive performance of layers when it was fortified with certain EAA, though poor albumen quality, and ileal inflammation were occurred. The addition of Ser to LCP diets improved performance probably through enhancing humoral and ileal mucosal immunity and attenuating the ileal inflammation of layers.

Effects of Selenium Conjugated to Insect Protein on Pharmacokinetics of Florfenicol and Enrofloxacin in Laying Hens.

In the context of increasing awareness on the dietary supplementation of organic selenium in commercial poultry production and ensuring safe egg production, the present study investigated the effects of selenium on the pharmacokinetics of the therapeutic use of florfenicol and enrofloxacin from perspectives of laying performance, selenium deposition in eggs, and drug residue in plasma, organs, and eggs. A 2 × 3 factorial arrangement with two kinds of drugs (florfenicol vs. enrofloxacin, 200 mg/kg) and three levels of dietary organic selenium SCIP (selenium conjugated to insect protein) (0, 2, and 5 mg/kg) was designed together with a blank control group. Healthy Hy-Line Brown laying hens (n = 252, 40-week-old and 90.0 ± 1.7% of egg production rate) were randomly allocated into one of seven treatments with six replicates and six hens per replicate. The experiment lasted for 42 days and consisted of three periods (adjusted stage, depositional stage, and eliminating stage) of 14 days each. These stages entail feeding of the laying hens with basal diets, addition of drugs and selenium synchronously into the diets, drug withdrawal from diet, and supply of selenium uninterruptedly in the diet. Egg production and feed intake were recorded on daily and weekly bases, respectively. The selenium content in egg yolk, egg white, and whole eggs and the drug residues in eggs, plasma, liver, kidney, and breast muscle were determined on days 2, 3, 5, 6, 7, 9, 11, and 14 of the depositional and eliminating stages. There was no significant difference (p > 0.05) in egg production among the dietary treatments, but feed intake decreased significantly (p < 0.05) in the drug treatment group compared to other groups. Dietary organic selenium decreased the residue of drugs in tissues and eggs, while the metabolism and deposition of selenium in laying hens were suppressed due to drug effects. The results of the present study are of significance to enrich the knowledge of the pharmacokinetics of florfenicol and enrofloxacin in laying hens and ensure the quality of poultry products.

The partial replacement of sodium chloride with sodium bicarbonate or sodium sulfate in laying hen diets improved laying performance, and eggshell quality and ultrastructure.

This study investigated the effects of dietary chloride (Cl) reduction on laying performance and eggshell quality by substitution of sodium bicarbonate (NaHCO3) or sodium sulfate (Na2SO4) for part of dietary sodium chloride (NaCl), and further explored its mechanism for improving eggshell quality. A total of 360 29-wk-old Hy-line Brown laying hens were randomly allocated to 5 dietary treatments, including a basal diet contained 0.33% NaCl (control group, 0.27% dietary Cl), and 4 experimental diets that contained 0.21% and 0.15% dietary Cl by substituting Na2SO4 or NaHCO3 for part of NaCl in the basal diet. No significant differences were observed in blood Na+, Cl-, K+ and Ca2+ levels and pH value as well as serum creatinine and uric acid contents among 5 treatments (P > 0.05). Dietary Cl reduction increased egg production and ADFI during wk 33 to 36, 37 to 40 and 29 to 40 of age and decreased feed conversion ratio during wk 37 to 40 of age (P < 0.05). The hens fed with diets containing 0.15% Cl increased eggshell breaking strength, thickness and weight ratio in wk 40 of age (P < 0.05). Birds fed with dietary 0.21% and 0.15% Cl exhibited higher effective layer thickness and lower mammillary layer thickness of eggshell than those fed with dietary 0.27% Cl (P < 0.05). Apparent Ca metabolizability of hens was increased with dietary Cl reduction (P < 0.05). Total Ca of eggshell of dietary 0.15% Cl group was higher than that of dietary 0.27% Cl group (P < 0.05). No significant differences in laying performance, eggshell quality and Ca metabolism of layers were observed between Na2SO4 and NaHCO3 replacement groups (P > 0.05). Overall, dietary Cl reductions could improve laying performance and eggshell quality by substitution of NaHCO3 or Na2SO4 for part of NaCl, and there were no differences in the improvements between these two substitutes. The improved eggshell quality may be attributed to improved eggshell ultrastructure and increased supply of eggshell Ca2CO3.

[Effect of combination of acupuncture and psychological intervention on attention, response inhibition and cerebral blood flow in children with attention deficit hyperactivity disorder].

OBJECTIVE: To observe the clinical curative effect on attention deficit hyperactivity disorder (ADHD) and explore the relevant mechanism of acupuncture in treatment. METHODS: A total of 100 ADHD children were randomized into an observation group (50 cases, 2 cases dropped off) and a control group (50 cases, 1 case dropped off). In the control group, the routine psychological intervention was used. In the observation group, on the base of the treatment as the control group, acupuncture was applied to Taichong (LR 3), Neiguan (PC 6), Shenmen (HT 7), Sanyinjiao (SP 6), Baihui (GV 20), Sishencong (EX-HN 1), etc., once daily, for 3 months. The Cambridge neuropsychological tests automated battery (CANTAB) was adopted to evaluate attention and response inhibition in two groups before and after treatment. Digi-Lite color transcranial Doppler was used to measure cerebral arterial blood velocity. The therapeutic effect was compared between the two groups. RESULTS: Regarding evaluation of attention, the mean delay time in the observation group after treatment was shorter than that before treatment and that in the control group separately (P<0.05), and rapid visual information processing A' statistics (RVP A') value was higher than that before treatment and that in the control group separately (P<0.05). For evaluation of response inhibition, the number of Go signal error, the time of stop signal response and the number of stop Go signal error after treatment were all reduced as compared with those before treatment in the two groups (P<0.05). The number of stop signal error was also reduced after treatment as compared with that before treatment in the observation group (P<0.05). The number of Go signal error and the number of stop signal error in the observation were lower than those in the control group (P<0.05). After treatment, the average flow velocity of bilateral posterior cerebral artery (PCA) was increased than that before treatment in the two groups (P<0.05). The average flow velocity of the left middle cerebral artery (MCA-L) and bilateral anterior cerebral artery (ACA) after treatment was increased than that before treatment in the observation group (P<0.05), and the average flow velocity of left PCA, MCA-L and bilateral ACA in the observation group was faster than that in the control group (P<0.05). The total effective rate was 90.5% (19/21) in the observation group, remarkably higher than 50.0% (11/22) in the control group in children with attention deficit (P<0.05). The total effective rate was 83.3% (40/48) in the observation group, also higher than 63.3% (31/49) in the control group (P<0.05). CONCLUSION: Acupuncture combined with psychological intervention may improve attention and response inhibition in ADHD children, which is possibly related to the regulation of cerebral blood flow.

Effects of dietary Bacillus subtilis supplementation and calcium levels on performance and eggshell quality of laying hens in the late phase of production.

The objective of this study was to evaluate the effects of dietary Bacillus subtilis supplementation and calcium (Ca) levels on performance, eggshell quality, intestinal morphology, and relative calbindin-D28k (CALB1) mRNA level of laying hens in the late phase of production. An experiment employing a 2 × 3 factorial arrangement of 3 levels of Ca (3.5, 4.0, and 4.5%) and the absence or presence of B. subtilis was carried out with a total of 576 Hy-Line Brown laying hens aged 72 to 79 wk. Every group had 8 replicates of 12 birds each. The results showed that 4.0 and 4.5% Ca levels improved (P < 0.05) apparent retention and serum Ca content of aged laying hens. Compared with the 3.5% Ca level, the 4.0% Ca level in diets increased (P < 0.05) thickness, eggshell weight, shell ratio, and eggshell Ca content of aged laying hens. Moreover, breaking strength, thickness, eggshell weight, shell ratio, eggshell Ca content, apparent retention of Ca in g/day, apparent retention of Ca in percent, villus height, villus height/crypt depth, serum Ca level, and relative CALB1 mRNA level of aged laying hens were all increased (P < 0.05) by B. subtilis supplementation in diets. The supplemental B. subtilis decreased feed conversion ratio (P = 0.001) significantly. In addition, there was an interaction effect between increased Ca levels from 3.5 to 4.5% and B. subtilis supplementation on crypt depth in the duodenum (P < 0.05). In conclusion, we found that both the increase in dietary Ca level from 3.5 to 4.5% and B. subtilis supplementation could enhance intestinal Ca absorption and improve eggshell quality of laying hens in the late phase of production (72-79 wk of age). Dietary supplementation of B. subtilis accompanying the 4.0% Ca level was appropriate in enhancement of eggshell quality.

Cyst(e)ine fortification in low crude protein diet improves growth performance of broilers by modulating serum metabolite profile.

This study was aimed to explore the metabolomical mechanisms for the potentially ameliorative effect of cyst(e)ine (Cys) fortification on growth performance of broilers fed low crude protein (CP) diet. A total of 432 1-d-old broilers were randomly divided into 6 groups, each of which received one of the following diets: normal-CP diet (positive control, PC), low-CP diet (negative control, NC), NC diet fortified with 0.05%, 0.1%, 0.15% or 0.2% of Cys. Samples were collected on d 42. Results showed that increasing Cys fortification quadratically elevated (P < 0.05) the accumulative growth performance and leg muscle yield of broilers fed NC diet, with 0.1% being the optimal dose. Thus, samples from PC, NC and NC plus 0.1% Cys (NCC) groups were selected for further analysis. Both dietary CP reduction and fortification of 0.1% Cys in NC diet caused complex changes (P < 0.05) in serum amino acids and some other metabolites primarily involved in lipid metabolism. Multiple lipogenesis-related pathways were regulated (P < 0.05) following Cys fortification in NC diet, which could at least partially interpret the benefit of Cys fortification in NC diet on broiler performance. In conclusion, fortifying low-CP diet with 0.1% Cys promoted the growth performance of broilers probably through modulating serum metabolite profile.

[Effect of maternal immune level at different pregnancy stages on cow's milk protein allergy in infants].

OBJECTIVE: To study the association between maternal Th1/Th2 immune level at different pregnancy stages and cow's milk protein allergy (CMPA) in infants. METHODS: The healthy women with a singleton pregnancy, as well as their offspring, who attended Yidu Central Hospital of Weifang and Qingzhou Traditional Chinese Medicine Hospital from July 2016 to December 2018 were enrolled. The maternal levels of interleukin-2 (IL-2), interferon gamma (IFN-γ), interleukin-4 (IL-4), and interleukin-10 (IL-10) at the second and third trimesters of pregnancy were measured. A CMPA questionnaire survey was conducted within one year after birth. Food avoidance and cow's milk oral challenge tests were performed in infants suspected of CMPA. The 48 infants who met the diagnostic criteria for CMPA were included in the observation group, and the remaining 977 normal infants were included in the control group. A univariate analysis was performed on the infants with CMPA. A Poisson regression analysis was used to determine the association between maternal Th1/Th2 immune factors at different pregnancy stages and CMPA. RESULTS: The detection rate of CMPA was 4.68%. The clinical manifestations included the symptoms of the digestive system, skin, and respiratory system and other symptoms. The univariate analysis showed that compared with the control group, the observation group had significantly higher incidence rates of maternal food allergy and maternal history of allergic diseases (P<0.05) and a significantly lower breastfeeding rate (P<0.05). The observation group had significantly lower maternal levels of IL-2 (second and third trimesters) and IFN-γ (third trimester) than the control group (P<0.05). Maternal low IFN-γ at the third trimester and maternal low IL-2 at the second and third trimesters were significantly associated with CMPA in infants (P<0.05). After correction of the factors of breastfeeding, maternal food allergy, and maternal history of allergic diseases, it was found that maternal low IL-2 and IFN-γ at the third trimester were still significantly associated with CMPA in infants (P<0.05). CONCLUSIONS: The maternal decrease in Th1 level at the third trimester of pregnancy may lead to the change in fetal immunity and thus increase the risk of CMPA in offspring.

The Response of Broiler Chickens to Dietary Soybean Meal Reduction with Glycine and Cysteine Inclusion at Marginal Sulfur Amino Acids (SAA) Deficiency.

The responses of broiler chickens to dietary protein reduction were investigated in the presence of glycine and cysteine inclusion at the marginal deficiency of sulfur-containing amino acids. A total of 432 broiler chickens were allotted to six dietary treatments; SP1 is standard protein diet with 100% total sulfur amino acids (TSAA), SP2 is standard protein diet with 85% TSAA, RP is reduced protein diet without glycine and cysteine supplementation, RPC is reduced protein diet with cysteine supplementation at 0.1%, and RPG is reduced protein diet with 1% glycine supplementation, while RPGC is reduced protein diet with 0.1% cysteine and 1% glycine supplementation. In this study, 4.5% protein is reduced in diets-thus, 17.5% CP (crude protein) for starter phase and 15.5% CP for the grower phase. Reduced protein diets contained 85% TSAA. Broiler chickens fed standard protein diet SP2 had superior bodyweight (BW) (p ≤ 0.05) in the starter and grower phase, average daily gain (ADG) (p ≤ 0.05) in the starter and entire feeding period, average daily feed intake (ADFI) (p ≤ 0.05) in the starter phase, and better feed conversion ratio (FCR) (p ≤ 0.05) in the starter, grower and entire feeding period; however, RPGC showed higher ADG (p ≤ 0.05) in the grower phase, and ADFI (p ≤ 0.05) in the grower and entire feeding period. RPC and RPG diet improved BW (p ≤ 0.05), ADG (p ≤ 0.05), ADFI (p ≤ 0.05), and better FCR (p ≤ 0.05) in starter, grower, entire feeding period compared to RP. The RPGC group had higher BW (p ≤ 0.05), ADG (p ≤ 0.05), ADFI (p ≤ 0.05) and better FCR (p ≤ 0.05) compared to the RPC group. Blood biochemical parameters showed that Broiler chickens fed on the SP2 diet had higher levels of total protein (TP) (p ≤ 0.05), albumin (ALB) (p ≤ 0.05), creatinine (CRE) (p ≤ 0.05), and aspartate aminotransferase (AST) (p ≤ 0.05) and, lower level of uric acid (UA) (p ≤ 0.05), blood urea nitrogen (BUN) (p ≤ 0.05), glucose (GLU) (p ≤ 0.05), and alanine aminotransferase (ALT) (p ≤ 0.05) in the starter phase; however, higher level of TP (p ≤ 0.05), GLU (p ≤ 0.05), CRE (p ≤ 0.05), and AST (p ≤ 0.05), and lower level of ALB (p ≤ 0.05), UA (p ≤ 0.05), and ALT (p ≤ 0.05) in the grower phase; RPGC had higher level of TP (p ≤ 0.05), UA (p ≤ 0.05), GLU (p ≤ 0.05), ALT (p ≤ 0.05) and AST (p ≤ 0.05), and lower level of ALB (p ≤ 0.05), BUN (p ≤ 0.05), and CRE (p ≤ 0.05) in the starter phase; however, in grower phase, RPGC had higher level of TP (p ≤ 0.05), and ALB (p ≤ 0.05), and lower level of UA (p ≤ 0.05), CRE (p ≤ 0.05), ALT (p ≤ 0.05), and AST (p ≤ 0.05). Free amino acids profile showed that broiler fed on standard protein diet SP2 had reduced the methionine (p ≤ 0.05) concentration; RPC increased the concentrations of taurine (p ≤ 0.05), phosphoethanolamine (p ≤ 0.05), threonine (p ≤ 0.05), valine (p ≤ 0.05), isoleucine (p ≤ 0.05), phenylalanine (p ≤ 0.05), ornithine (p ≤ 0.05), and lysine (p ≤ 0.05) and reduced the citrulline (p ≤ 0.05) concentration; RPG increased the concentration of glutamate (p ≤ 0.05), glycine (p ≤ 0.05), cysteine (p ≤ 0.05), and arginine (p ≤ 0.05), and decreased the concentration of tyrosine (p ≤ 0.05); and RPGC increased the concentration of serine (p ≤ 0.05) and reduced the concentration of hydroxyproline (p ≤ 0.05). Serum metabolites analysis showed that reduced protein downregulated the 54 metabolites; however, glycine fortification up-regulated the Benzamide, Pro-Ser, N-Carbamylglutamate, D-gluconate, and Gamma-Glutamylcysteine. Carcass quality showed that SP2 decreased the abdominal fat percentage (p ≤ 0.05). Nitrogen digestibility was higher by the diet RP (p ≤ 0.05). This study demonstrated that protein content could be reduced up to 4.5% with 1% glycine and 0.1% cysteine fortification in diet, which has the potential to inhibit the adverse effect of reduced protein and attain the standard growth performance.

Dynamic alterations in early intestinal development, microbiota and metabolome induced by in ovo feeding of L-arginine in a layer chick model.

BACKGROUND: Prenatal nutrition is crucial for embryonic development and neonatal growth, and has the potential to be a main determinant of life-long health. In the present study, we used a layer chick model to investigate the effects of in ovo feeding (IOF) of L-arginine (Arg) on growth, intestinal development, intestinal microbiota and metabolism. The treatments included the non-injected control, saline-injected control, and saline containing 2, 6, or 10 mg Arg groups. RESULTS: IOF Arg increased early intestinal index and villus height, and enhanced uptake of residual yolk lipid, contributing to subsequent improvement in the early growth performance of chicks. Prenatal Arg supplementation also increased the early microbial α-diversity, the relative abundance of Lactobacillales and Clostridiales, and decreased the relative abundance of Proteobacteria of cecum in chicks. Furthermore, the shift of cecal microbiota composition and the colonization of potential probiotics were accelerated by IOF of Arg. Simultaneously, metabolomics showed that metabolisms of galactose, taurine-conjugated bile acids and lipids were modulated to direct more energy and nutrients towards rapid growth of intestine at the beginning of post-hatch when embryos received IOF of Arg. CONCLUSIONS: Prenatal Arg supplementation showed beneficial effects on the early intestinal development, cecal microbiota and host metabolism of layer chicks, contributing to subsequent improvement in the early growth performance. These findings provide new insight into the role of IOF of Arg in the establishment of the gut microbiota of newly-hatched layer chicks, and can expand our fundamental knowledge about prenatal nutrition, early bacterial colonization and intestinal development in neonate.

Comparative effects of dietary microalgae oil and fish oil on fatty acid composition and sensory quality of table eggs.

This study was conducted to investigate the comparative effects of dietary supplementation with microalgae oil or fish oil on fatty acid composition, sensory quality, and overall acceptability of table eggs. A total of six hundred thirty, 30-week-old, Hy-Line Brown laying hens were allocated to 7 dietary treatments. Layers were fed with a control diet or the control diet supplemented with graded levels of docosahexaenoic acid (DHA; 1.25, 2.50, and 5.00 mg/g feed) derived from microalgae oil or fish oil. The feeding trial lasted 10 wk. Enrichment of eggs with DHA by dietary supplementation with microalgae oil or fish oil enhanced yolk DHA and total n-3 polyunsaturated fatty acid (PUFA) enrichment and reduced n-6-to-n-3 ratio in a dose-response manner, whereas the efficiency decreased (P < 0.05). The overall efficiency of DHA incorporation into eggs was similar for the 2 sources (P > 0.05). In comparison with fish oil, dietary microalgae oil supplementation resulted in higher scores for egg flavor and overall acceptability, both of which declined linearly in response to DHA supplementation (P < 0.05). Among the aroma and flavor characteristics, fishy aroma and flavor scores increased linearly and quadratically (P < 0.05) in response to dietary DHA supplementation, and egg aroma and flavor and milky flavor scores decreased linearly (P < 0.05). The results from partial least squares analysis showed that fishy flavor and aroma were closely associated with DHA, α-linolenic acid, and total n-3 PUFA, and oleic acid, arachidonic acid, and dihomo-gamma-linolenic acid were more related to egg aroma and flavor. The results suggested that microalgae oil would be more promising for egg DHA enrichment owing to better sensory quality of the resultant eggs.

Standardized ileal digestible amino acid and metabolizable energy content of wheat from different origins and the effect of exogenous xylanase on their determination in broilers.

This study was conducted to determine the standardized ileal digestible amino acids (SID AA) and nitrogen-corrected apparent metabolizable energy (AMEn) contents of 6 wheats from different origins in China and incidentally to investigate the effects of exogenous xylanase addition on SID AA and AMEn determination in broiler chicks. A total of 480 chicks were divided into 48 cages of 10 birds each balanced for body weight and fed 8 types of diets in a completely randomized design (6 replicated cages per diet) from 21 to 26 d of age. The individual wheat constituted the only source of crude protein in a semi-purified experimental diet. A nitrogen-free diet was designed to estimate basal endogenous AA loss and determine the SID AA. Titanium oxide (0.3%) was used as an indigestibility marker, and nutrient digestibility and retention were determined by the substitution method. From day 24 to 26, excreta samples were collected for AMEn determination. On day 26, the birds were euthanized, and ileum contents were obtained for AA digestibility determination. Wheat from Gansu had greater (P < 0.05) SID AA contents except Lys, Thr, Phe, and Cys, with a higher (P < 0.001) AMEn (11.83 MJ/kg) than the other wheats. The SID content of mean indispensable amino acids and dispensable amino acids were 87.35% and 88.17%, respectively, and the average AMEn value of 6 wheats was 11.14 MJ/kg. Compared with the diet without xylanase, the added xylanase resulted in higher (P < 0.05) SID contents of Met, Lys, Trp, Arg, Ile, Leu, Val, Gly, Asp, Glu, Pro, and Ala; the SID AA values were raised by 1.96% (mean of all AA); and the AMEn content was significantly increased (+0.87 MJ/kg) (P < 0.05). In conclusion, origins of wheats have significant effects on SID AA and AMEn values which were positively correlated with crude protein content of wheat; exogenous xylanase addition to a wheat-based poultry diet could significantly improve SID AA and AMEn contents for broilers.

Effect of in ovo feeding of ß-hydroxy-ß-methylbutyrate on hatchability, muscle growth and performance in prenatal and posthatch broilers.

BACKGROUND: ß-Hydroxy-ß-methylbutyrate (HMB) is the metabolite of leucine that plays an important role in muscle protein metabolism. The objective of the present study was to determine the effects of in ovo feeding (IOF) of HMB at 7 days of incubation (DOI) via air cell or 18 DOI via amnion on hatchability, muscle growth and performance in prenatal and posthatch broilers. RESULTS: IOF of HMB via air cell at 7 DOI increased hatchability by 4.34% compared with the control (89.67% versus 85.33%). Birds in IOF groups exhibited higher body weight, average daily body weight gain and pectoral muscle percentage. Furthermore, IOF of HMB significantly increased the level of plasma growth hormone, insulin and insulin-like growth factor-1. Chicks hatched from IOF treatment had larger diameters of muscle fiber and higher mitotic activity of satellite cells at early posthatch age. IOF of HMB activated satellite cells by upregulation of mRNA expression of myogenic transcription factors, myogenic differentiation one (MyoD) and myogenin. Chicks hatched from air cell injection group had higher pectoral muscle percentage at 5 d posthatch and greater satellite cell mitotic activity at 7 d posthatch than counterparts from amnion injection group. CONCLUSIONS: IOF of HMB via amnion at 18 DOI or especially via air cell at 7 DOI could be used as an effective approach to enhance hatchability, productive performance and breast muscle yield in broilers. © 2019 Society of Chemical Industry.

[Acupuncture combined with medication improves endocrine hormone levels and ovarian reserve function in poor ovarian response patients undergoing in vitro fertilization-embryo transplantation].

OBJECTIVE: To observe the clinical effect of acupuncture plus medication in the treatment of poor ovarian response (POR) patients and to explore its mechanisms in assisting pregnancy. METHODS: A total of 100 volunteer POR women undergoing in vitro fertilization-embryo transplantation (IVF-ET) were recruited in the present study. On the 1st cycle of IVF-ET, these POR women received microstimulation of ovulation program (oral administration of Clomiphene, muscular injection of Menotrophin, Chorionic Gonadotrophin triggering, etc.). Before receiving the 2nd period of IVF-ET, these patients were equally and randomly divided into control, medication (Climen, composed of estradiol valerate and cyproterone acetate), acupuncture and acupuncture+medication (combined treatment) groups according to the random number table. Patients of the medication group were asked to orally take Climen (1 tablet/d for 21 days) beginning from the 3rd day of the menstruation, which was repeated for 3 menstrual cycles. Patients of the acupuncture group received manual acupuncture stimulation of Guanyuan (CV4), and bilateral Taixi (KI3), Sanyinjiao (SP6) of and Tai-chong (LR3) from day 8 to 15 of menstruation (follicular phase), once daily for 3 menstrual cycles. On the 2nd day of menstruation of the 1st and 2nd IVF-ET cycle, the ovarian reserve function was detected, including measurement of serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) contents by using radioimmunoassay, and serum anti-mullerian hormone (AMH) level by using ELISA, and the antral follicles count (AFC) of the ovaries by using a color Doppler ultrasonic diagnosis apparatus. At the end of ovulation induction, the assisted pregnancy indexes and outcomes were detected, including administration of dosage and days of gonadotropin (Gn), the diameter of dominant oocyte, level of E2 on the trigger day, the numbers of ultrasound-guided-retrieved oocyte and the cultivated high-quality embryo (grade 1 and 2). RESULTS: After the treatment, the contents of serum FSH, LH and E2 in the medication, acupuncture and combined treatment groups were significantly reduced (P<0.05), and the content of serum AMH and ovary AFC in the medication, acupuncture and combined treatment groups were obviously increased (P<0.05) in comparison with their own pre-treatment (P<0.05). Compared with the control group, the levels of FSH, LH and E2 contents were notably decreased, and the AMH content and AFC significantly increased in the medication, acupuncture and combined treatment groups (P<0.05). Comparison among the three treatment groups showed that the levels of FSH, LH and E2 were notably lower in the combined treatment group than in the acupuncture group (P<0.05), and the AFC was markedly more in the combined treatment group than in the acupuncture group (P<0.05), and LH content was also significantly lower in the combined treatment group than in the medication group (P<0.05). Regarding the pregnancy-assistant indexes and outcomes, the dosage of Gn in the combined treatment group and the number of Gn administration days in the acupuncture and combined treatment groups were significantly fewer (P<0.05), serum E2 levels on the trigger day in the medication, acupuncture and combined treatment groups, and the high-quality embryo number in the combined treatment group were considerably higher in comparison with their own levels of the 1st IVF-ET cycle in each group (P<0.05). Comparison among the three groups showed that the dosage and number of administration days of Gn in the combined treatment group were fewer than those in the medication group(P<0.05), and the dosage of Gn in the combined treatment group were significantly lower than that in the acupuncture group (P<0.05), E2 content of the combined treatment group was markedly higher than that of the medication and acupuncture groups (P<0.05). The high-quality embryo number in the combined treatment group was obviously larger than that of the medication group (P<0.05). No significant differences were found between the 2nd and 1st IVF-ET cycles in the number of the obtained oocyte, between the acupuncture and medication groups in the E2 content at the trigger day, and the numbers of Gn administration days and the high-quality embryo (P>0.05). CONCLUSION: Acupuncture combined with medication improves the level of endocrinal hormones and ovarian reservation function in POR women undergoing IVF-ET, benefiting the ovary environment of pregnancy.

Supplemental Plant Extracts From Flos lonicerae in Combination With Baikal skullcap Attenuate Intestinal Disruption and Modulate Gut Microbiota in Laying Hens Challenged by Salmonella pullorum.

Dietary inclusions of baicalin and chlorogenic acid were beneficial for intestinal health in pigs. Nevertheless, it is unknown whether these plant-derived products had protection for intestine against bacterial challenge in chickens. This study was aimed at evaluating the potential mitigating effects of plant extracts (PE) from Flos lonicerae combined with Baikal skullcap (the active components are chlorogenic acid and baicalin) on intestinal disruption and dysbacteriosis induced by Salmonella pullorum in laying hens. A total of 216 41-week-old layers were randomly divided into 3 groups (6 replicates per group): negative control (NC), S. pullorum-infected positive control (PC), and the S. pullorum-infected group with supplementation of PE at 1000 mg/kg. All birds except those in NC were challenged with S. pullorum at the end of 4 weeks of the experiment. S. pullorum challenge impaired (P < 0.05) the production performance (egg production, feed intake, and feed efficiency) of laying hens, increased (P < 0.05) serum endotoxin content and frequency of Salmonella-positive organs, as well as up-regulated (P < 0.05) ileal expression of pro-inflammatory cytokines including IFNG, TNFA, IL8, and IL1B, whereas PE addition reversed (P < 0.05) these changes and increased (P < 0.05) ileal IL10 expression. Supplemental PE moderated ileal microbiota dysbiosis in challenged birds, characterized by a reduced abundance of Firmicutes along with increased abundances of Bacteroidetes (Bacteroides), Deferribacteres and several butyrate-producers such as Prevotellaceae, Faecalibacterium, Blautia, Butyricicoccus, Lachnoclostridium, and Olsenella, which may assist with energy harvesting and boost anti-inflammatory capacity of host. The decreased abundance of Firmicutes with the increased abundance of Bacteroidetes caused by PE addition had positive correlations with the decreased expression of ileal pro-inflammatory cytokines. The increased abundances of Bacteroidetes (Bacteroides) and Prevotellaceae following PE addition were also positively correlated with the improvement of performance (egg production and feed intake) of laying hens. Collectively, supplemental PE from Flos lonicerae in combination with Baikal skullcap alleviated S. pullorum-induced intestinal disruption and performance impairment in laying hens, which could be at least partially responsible by the modulation of gut microbial composition.

Conjugated linoleic acid regulates lipid metabolism through the expression of selected hepatic genes in laying hens.

The effect of dietary conjugated linoleic acid (CLA) supplementation on lipid metabolism in laying hens was investigated. A total of 360 eighteen-wk-old Hy-Line Brown layers were randomly divided into 4 groups that consisted of 6 replicates with 15 birds each. Birds were fed basal diets with 0, 1%, 2%, and 4% CLA addition. The experiment lasted for 56 D after a 7-D adaptation period. Results showed that dietary CLA addition linearly reduced (P < 0.05) abdominal fat percentage but linearly increased (P < 0.05) relative liver weight of layers on day 56. A linear reduction (P < 0.05) in serum low-density lipoprotein cholesterol (LDL-C) level and a linear elevation (P < 0.05) in the ratio of serum high-density lipoprotein cholesterol level to LDL-C level of layers on both days 28 and 56 were observed with dietary CLA addition, which also linearly decreased (P < 0.05) cholesterol content in the liver of layers on day 56 as well as in eggs on both days 28 and 56. Besides, there were linear reductions (P < 0.05) in the gene expression and contents of hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) and cholesterol 7 alpha hydroxylase 1 (CYP7A1), along with a linear increase (P < 0.05) in the gene expression and content of hepatic low-density lipoprotein receptor (LDLR) in layers responded to dietary CLA addition. In conclusion, dietary CLA supplementation decreased the accumulation of lipids including abdominal fat and cholesterol in the liver and egg of laying hens, probably by upregulating hepatic LDLR expression and downregulating hepatic HMGR and CYP7A1 expression.

Trans10, cis12-conjugated linoleic acid exhibits a stronger antioxidant capacity than cis9, trans11-conjugated linoleic acid in primary cultures of laying hen hepatocytes.

The objective of this study consisting of 2 trials was to investigate the antioxidant role of conjugated linoleic acid (CLA) isomers (c9, t11-CLA and t10, c12-CLA) and the underlying mechanism by which they act in modulating redox status in a primary laying hen hepatocyte culture. In trial 1, the cytotoxicity of CLA isomers or linoleic acid (LA) (0, 25, 50, 100, 200, 400, 800 µmol/L) was evaluated by 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide (MTT) assay. The concentration of CLA isomers or LA (25, 50, 100 µmol/L) for proper antioxidant activity was evaluated by measuring the antioxidant enzyme activity. In trial 2, there were 5 groups: control group, cells were untreated; H2O2 group, cells were exposed to 4 mmol/L H2O2 for 2 h; c9, t11 or t10, c12 or LA group, cells were treated with c9, t11-CLA or t10, c12-CLA or LA for 24 h and then exposed to 4 mmol/L H2O2 for 2 h. Trial 1 showed that the non-toxic dose range for CLA isomers was 0 to 200 µmol/L. The optimum concentration of c9, t11-CLA and t10, c12-CLA for trial 2 was 100 µmol/L. In trial 2, pretreatment with t10, c12-CLA but not c9, t11-CLA attenuated the increase in reactive oxygen species (ROS) compared to hydrogen peroxide (H2O2) group (P < 0.05). t10, c12-CLA elevated the superoxide dismutase (SOD) and catalase (CAT) activities compared with the H2O2 group (P < 0.05). In addition, t10, c12-CLA up-regulated the mRNA expression of nuclear factor E2-related factor-2 (Nrf2) as well as its target genes, Cu-Zn superoxide dismutase (SOD1) and CAT (P < 0.05). Pretreatment with t10, c12-CLA but not c9, t11-CLA decreased Nrf2 protein expression in the cytoplasm and increased Nrf2 protein expression in the nucleus compared with the H2O2 group (P < 0.05). The results indicate that t10, c12-CLA exhibits a stronger antioxidant capacity than c9, t11-CLA in primary cultured laying hen hepatocytes. t10, c12-CLA increases the activity and mRNA expression of antioxidant enzymes via facilitating nuclear translocation of Nrf2.